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At least 1,477 records · Page 82
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DOE OSTI2013

Genome Variations Associated with Viral Susceptibility and Calcification in Emiliania huxleyi

Emiliania huxleyi, a key player in the global carbon cycle is one of the best studied coccolithophores with respect to biogeochemical cycles, climatology, and host-virus interactions. Strains of E. huxleyi show phenotypic plasticity regarding growth behaviour, light-response, calcification, acidification, and virus susceptibility. This phenomenon is likely a consequence of genomic differences, or transcriptomic responses, to environmental conditions or threats such as viral infections. We used an E. huxleyi genome microarray based on the sequenced strain CCMP1516 (reference strain) to perform comparative genomic hybridizations (CGH) of 16 E. huxleyi strains of different geographic origin. We investigated the genomic diversity and plasticity and focused on the identification of genes related to virus susceptibility and coccolith production (calcification). Among the tested 31940 gene models a core genome of 14628 genes was identified by hybridization among 16 E. huxleyi strains. 224 probes were characterized as specific for the reference strain CCMP1516. Compared to the sequenced E. huxleyi strain CCMP1516 variation in gene content of up to 30 percent among strains was observed. Comparison of core and non-core transcripts sets in terms of annotated functions reveals a broad, almost equal functional coverage over all KOG-categories of both transcript sets within the whole annotated genome. Within the variable (non-core) genome we identified genes associated with virus susceptibility and calcification. Genes associated with virus susceptibility include a Bax inhibitor-1 protein, three LRR receptor-like protein kinases, and mitogen-activated protein kinase. Our list of transcripts associated with coccolith production will stimulate further research, e.g. by genetic manipulation. In particular, the V-type proton ATPase 16 kDa proteolipid subunit is proposed to be a plausible target gene for further calcification studies.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

Development of an Alfalfa SNP Array and Its Use to Evaluate Patterns of Population Structure and Linkage Disequilibrium

A large set of genome-wide markers and a high-throughput genotyping platform can facilitate the genetic dissection of complex traits and accelerate molecular breeding applications. Previously, we identified about 0.9 million SNP markers by sequencing transcriptomes of 27 diverse alfalfa genotypes. From this SNP set, we developed an Illumina Infinium array containing 9,277 SNPs. Using this array, we genotyped 280 diverse alfalfa genotypes and several genotypes from related species. About 81% (7,476) of the SNPs met the criteria for quality control and showed polymorphisms. The alfalfa SNP array also showed a high level of transferability for several closely related Medicago species. Principal component analysis and model-based clustering showed clear population structure corresponding to subspecies and ploidy levels. Within cultivated tetraploid alfalfa, genotypes from dormant and nondormant cultivars were largely assigned to different clusters; genotypes from semidormant cultivars were split between the groups. The extent of linkage disequilibrium (LD) across all genotypes rapidly decayed to 26 Kbp at r 2 = 0.2, but the rate varied across ploidy levels and subspecies. A high level of consistency in LD was found between and within the two subpopulations of cultivated dormant and nondormant alfalfa suggesting that genome-wide association studies (GWAS) and genomic selection (GS) could be conducted using alfalfa genotypes from throughout the fall dormancy spectrum. However, the relatively low LD levels would require a large number of markers to fully saturate the genome.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

Roles of Gibberellin Catabolism and Signaling in Growth and Physiological Response to Drought and Short-Day Photoperiods in Populus Trees

Survival and productivity of perennial plants in temperate zones are dependent on robust responses to prolonged and seasonal cycles of unfavorable conditions. Here we report whole-genome microarray, expression, physiological, and transgenic evidence in hybrid poplar (Populus tremula × Populus alba) showing that gibberellin (GA) catabolism and repressive signaling mediates shoot growth inhibition and physiological adaptation in response to drought and short-day (SD) induced bud dormancy. Both water deprivation and SDs elicited activation of a suite of poplar GA2ox and DELLA encoding genes. Poplar transgenics with up-regulated GA 2-oxidase (GA2ox) and DELLA domain proteins showed hypersensitive growth inhibition in response to both drought and SDs. In addition, the transgenic plants displayed greater drought resistance as evidenced by increased pigment concentrations (chlorophyll and carotenoid) and reductions in electrolyte leakage (EL). Comparative transcriptome analysis using whole-genome microarray showed that the GA-deficiency and GA-insensitivity, SD-induced dormancy, and drought response in poplar share a common regulon of 684 differentially-expressed genes, which suggest GA metabolism and signaling plays a role in plant physiological adaptations in response to alterations in environmental factors. Our results demonstrate that GA catabolism and repressive signaling represents a major route for control of growth and physiological adaptation in response to immediate or imminent adverse conditions.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

Analysis of Global Gene Expression in Brachypodium distachyon Reveals Extensive Network Plasticity in Response to Abiotic Stress

Brachypodium distachyon is a close relative of many important cereal crops. Abiotic stress tolerance has a significant impact on productivity of agriculturally important food and feedstock crops. Analysis of the transcriptome of Brachypodium after chilling, high-salinity, drought, and heat stresses revealed diverse differential expression of many transcripts. Weighted Gene Co-Expression Network Analysis revealed 22 distinct gene modules with specific profiles of expression under each stress. Promoter analysis implicated short DNA sequences directly upstream of module members in the regulation of 21 of 22 modules. Functional analysis of module members revealed enrichment in functional terms for 10 of 22 network modules. Analysis of condition-specific correlations between differentially expressed gene pairs revealed extensive plasticity in the expression relationships of gene pairs. Photosynthesis, cell cycle, and cell wall expression modules were down-regulated by all abiotic stresses. Modules which were up-regulated by each abiotic stress fell into diverse and unique gene ontology GO categories. This study provides genomics resources and improves our understanding of abiotic stress responses of Brachypodium.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

Expression of a Humanized Viral 2A-Mediated lux Operon Efficiently Generates Autonomous Bioluminescence in Human Cells

Expression of autonomous bioluminescence from human cells was previously reported to be impossible, suggesting that all bioluminescent-based mammalian reporter systems must therefore require application of a potentially influential chemical substrate. While this was disproven when the bacterial luciferase (lux) cassette was demonstrated to function in a human cell, its expression required multiple genetic constructs, was functional in only a single cell type, and generated a significantly reduced signal compared to substrate-requiring systems. Here we investigate the use of a humanized, viral 2A-linked lux genetic architecture for the efficient introduction of an autobioluminescent phenotype across a variety of human cell lines. The lux cassette was codon optimized and assembled into a synthetic human expression operon using viral 2A elements as linker regions. Human kidney, breast cancer, and colorectal cancer cell lines were both transiently and stably transfected with the humanized operon and the resulting autobioluminescent phenotype was evaluated using common imaging instrumentation. Autobioluminescent cells were screened for cytotoxic effects resulting from lux expression and their utility as bioreporters was evaluated through the demonstration of repeated monitoring of single populations over a prolonged period using both a modified E-SCREEN assay for estrogen detection and a classical cytotoxic compound detection assay for the antibiotic Zeocin. Furthermore, the use of self-directed bioluminescent initiation in response to target detection was assessed to determine its amenability towards deployment as fully autonomous sensors. In all cases, bioluminescent measurements were supported with traditional genetic and transcriptomic evaluations. Our results demonstrate that the viral 2A-linked, humanized lux genetic architecture successfully produced autobioluminescent phenotypes in all cell lines tested without the induction of cytotoxicity. This autobioluminescent phenotype allowed for repeated interrogation of populations and self-directed control of bioluminescent activation with detection limits and EC 50 values similar to traditional reporter systems, making the autobioluminescent cells amenable to automated monitoring and significantly reducing the time and cost required to perform bioluminescent workflows.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

The Microbial Detection Array for Detection of Emerging Viruses in Clinical Samples - A Useful Panmicrobial Diagnostic Tool

Emerging viruses are usually endemic to tropical and sub-tropical regions of the world, but increased global travel, climate change and changes in lifestyle are believed to contribute to the spread of these viruses into new regions. Many of these viruses cause similar disease symptoms as other emerging viruses or common infections, making these unexpected pathogens difficult to diagnose. Broad-spectrum pathogen detection microarrays containing probes for all sequenced viruses and bacteria can provide rapid identification of viruses, guiding decisions about treatment and appropriate case management. We report a modified Whole Transcriptome Amplification (WTA) method that increases unbiased amplification, particular of RNA viruses. Using this modified WTA method, we tested the specificity and sensitivity of the Lawrence Livermore Microbial Detection Array (LLMDA) against a wide range of emerging viruses present in both nonclinical and clinical samples using two different microarray data analysis methods.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

Contrasting Metabolism in Perenniating Structures of Upland and Lowland Switchgrass Plants Late in the Growing Season

Background: Switchgrass (Panicum virgatum L.) is being developed as a bioenergy crop for many temperate regions of the world. One way to increase biomass yields is to move southern adapted lowland cultivars to more northern latitudes. However, many southerly adapted switchgrass germplasm can suffer significant winter kill in northerly climes. Materials and Methods: Here, we have applied next-generation sequencing in combination with biochemical analyses to query the metabolism of crowns and rhizomes obtained from two contrasting switchgrass cultivars. Crowns and rhizomes from field-grown lowland (cv Kanlow) and upland (cv Summer) switchgrass cultivars were collected from three randomly selected post-flowering plants. Summer plants were senescing, whereas Kanlow plants were not at this harvest date. Results: Principal component analysis (PCA) differentiated between both the Summer and Kanlow transcriptomes and metabolomes. Significant differences in transcript abundances were detected for 8,050 genes, including transcription factors such as WRKYs and those associated with phenylpropanoid biosynthesis. Gene-set enrichment analyses showed that a number of pathways were differentially up-regulated in the two populations. For both populations, protein levels and enzyme activities agreed well with transcript abundances for genes involved in the phenylpropanoid pathway that were upregulated in Kanlow crowns and rhizomes. The combination of these datasets suggests that dormancy-related mechanisms had been triggered in the crowns and rhizomes of the Summer plants, whereas the crowns and rhizomes of Kanlow plants had yet to enter dormancy. Conclusions: Delayed establishment of dormancy at more northerly latitudes could be one factor that reduces wintersurvival in the high-yielding Kanlow plants. Understanding the cellular signatures that accompany the transition to dormancy can be used in the future to select plants with improved winter hardiness.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

SPINE: SParse eIgengene NEtwork linking gene expression clusters in Dehalococcoides mccartyi to perturbations in experimental conditions

We present a statistical model designed to identify the effect of experimental perturbations on the aggregate behavior of the transcriptome expressed by the bacterium Dehalococcoides mccartyi strain 195. Strains of Dehalococcoides are used in sub-surface bioremediation applications because they organohalorespire tetrachloroethene and trichloroethene (common chlorinated solvents that contaminate the environment) to non-toxic ethene. However, the biochemical mechanism of this process remains incompletely described. Additionally, the response of Dehalococcoides to stress-inducing conditions that may be encountered at field-sites is not well understood. The constructed statistical model captured the aggregate behavior of gene expression phenotypes by modeling the distinct eigengenes of 100 transcript clusters, determining stable relationships among these clusters of gene transcripts with a sparse network-inference algorithm, and directly modeling the effect of changes in experimental conditions by constructing networks conditioned on the experimental state. Based on the model predictions, we discovered new response mechanisms for DMC, notably when the bacterium is exposed to solvent toxicity. The network identified a cluster containing thirteen gene transcripts directly connected to the solvent toxicity condition. Transcripts in this cluster include an iron-dependent regulator (DET0096-97) and a methylglyoxal synthase (DET0137). To validate these predictions, additional experiments were performed. Continuously fed cultures were exposed to saturating levels of tetrachloethene, thereby causing solvent toxicity, and transcripts that were predicted to be linked to solvent toxicity were monitored by quantitative reverse-transcription polymerase chain reaction. Twelve hours after being shocked with saturating levels of tetrachloroethene, the control transcripts (encoding for a key hydrogenase and the 16S rRNA) did not significantly change. By contrast, transcripts for DET0137 and DET0097 displayed a 46.8±11.5 and 14.6±9.3 fold up-regulation, respectively, supporting the model. This is the first study to identify transcripts in Dehalococcoides that potentially respond to tetrachloroethene solvent-toxicity conditions that may be encountered near contamination source zones in sub-surface environments.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

Widespread Polycistronic Transcripts in Fungi Revealed by Single-Molecule mRNA Sequencing

Genes in prokaryotic genomes are often arranged into clusters and co-transcribed into polycistronic RNAs. Isolated examples of polycistronic RNAs were also reported in some higher eukaryotes but their presence was generally considered rare. Here we developed a longread sequencing strategy to identify polycistronic transcripts in several mushroom forming fungal species including Plicaturopsis crispa, Phanerochaete chrysosporium, Trametes versicolor, and Gloeophyllum trabeum. We found genome-wide prevalence of polycistronic transcription in these Agaricomycetes, involving up to 8% of the transcribed genes. Unlike polycistronic mRNAs in prokaryotes, these co-transcribed genes are also independently transcribed. We show that polycistronic transcription may interfere with expression of the downstream tandem gene. Further comparative genomic analysis indicates that polycistronic transcription is conserved among a wide range of mushroom forming fungi. In summary, our study revealed, for the first time, the genome prevalence of polycistronic transcription in a phylogenetic range of higher fungi. Furthermore, we systematically show that our long-read sequencing approach and combined bioinformatics pipeline is a generic powerful tool for precise characterization of complex transcriptomes that enables identification of mRNA isoforms not recovered via short-read assembly.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

Strand-Specific RNA-Seq Analyses of Fruiting Body Development in Coprinopsis cinerea

We report that the basidiomycete fungus Coprinopsis cinerea is an important model system for multicellular development. Fruiting bodies of C. cinerea are typical mushrooms, which can be produced synchronously on defined media in the laboratory. To investigate the transcriptome in detail during fruiting body development, high-throughput sequencing (RNA-seq) was performed using cDNA libraries strand-specifically constructed from 13 points (stages/tissues) with two biological replicates. The reads were aligned to 14,245 predicted transcripts, and counted for forward and reverse transcripts. Differentially expressed genes (DEGs) between two adjacent points and between vegetative mycelium and each point were detected by Tag Count Comparison (TCC). To validate RNA-seq data, expression levels of selected genes were compared using RPKM values in RNA-seq data and qRT-PCR data, and DEGs detected in microarray data were examined in MA plots of RNA-seq data by TCC. We discuss events deduced from GO analysis of DEGs. In addition, we uncovered both transcription factor candidates and antisense transcripts that are likely to be involved in developmental regulation for fruiting.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2016

Functional and structural analysis of a highly-expressed Yersinia pestis small RNA following infection of cultured macrophages

Non-coding small RNAs (sRNAs) are found in practically all bacterial genomes and play important roles in regulating gene expression to impact bacterial metabolism, growth, and virulence. We performed transcriptomics analysis to identify sRNAs that are differentially expressed in Yersinia pestis that invaded the human macrophage cell line THP-1, compared to pathogens that remained extracellular in the presence of host. Using ultra high-throughput sequencing, we identified 37 novel and 143 previously known sRNAs in Y. pestis . In particular, the sRNA Ysr170 was highly expressed in intracellular Yersinia and exhibited a log2 fold change ~3.6 higher levels compared to extracellular bacteria. We found that knock-down of Ysr170 expression attenuated infection efficiency in cell culture and growth rate in response to different stressors. In addition, we applied selective 2’-hydroxyl acylation analyzed by primer extension (SHAPE) analysis to determine the secondary structure of Ysr170 and observed structural changes resulting from interactions with the aminoglycoside antibiotic gentamycin and the RNA chaperone Hfq. Interestingly, gentamicin stabilized helix 4 of Ysr170, which structurally resembles the native gentamicin 16S ribosomal binding site. Lastly, we modeled the tertiary structure of Ysr170 binding to gentamycin using RNA motif modeling. Integration of these experimental and structural methods can provide further insight into the design of small molecules that can inhibit function of sRNAs required for pathogen virulence.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

BIG LEAF is a regulator of organ size and adventitious root formation in poplar

Here we report the discovery through activation tagging and subsequent characterization of the BIG LEAF (BL) gene from poplar. In poplar, BL regulates leaf size via positively affecting cell proliferation. Up and downregulation of the gene led to increased and decreased leaf size, respectively, and these phenotypes corresponded to increased and decreased cell numbers. BL function encompasses the early stages of leaf development as native BL expression was specific to the shoot apical meristem and leaf primordia and was absent from the later stages of leaf development and other organs. Consistently, BL downregulation reduced leaf size at the earliest stages of leaf development. Ectopic expression in mature leaves resulted in continued growth most probably via sustained cell proliferation and thus the increased leaf size. In contrast to the positive effect on leaf growth, ectopic BL expression in stems interfered with and significantly reduced stem thickening, suggesting that BL is a highly specific activator of growth. Additionally, stem cuttings from BL overexpressing plants developed roots, whereas the wild type was difficult to root, demonstrating that BL is a positive regulator of adventitious rooting. Large transcriptomic changes in plants that overexpressed BL indicated that BL may have a broad integrative role, encompassing many genes linked to organ growth. Here, we conclude that BL plays a fundamental role in control of leaf size and thus may be a useful tool for modifying plant biomass productivity and adventitious rooting.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

Multi-time series RNA-seq analysis of Enterobacter lignolyticus SCF1 during growth in lignin-amended medium

The production of lignocellulosic-derived biofuels is a highly promising source of alternative energy, but it has been constrained by the lack of a microbial platform capable to efficiently degrade this recalcitrant material and cope with by-products that can be toxic to cells. Species that naturally grow in environments where carbon is mainly available as lignin are promising for finding new ways of removing the lignin that protects cellulose for improved conversion of lignin to fuel precursors. Enterobacter lignolyticus SCF1 is a facultative anaerobic Gammaproteobacteria isolated from tropical rain forest soil collected in El Yunque forest, Puerto Rico under anoxic growth conditions with lignin as sole carbon source. Whole transcriptome analysis of SCF1 during E.lignolyticus SCF1 lignin degradation was conducted on cells grown in the presence (0.1%, w/w) and the absence of lignin, where samples were taken at three different times during growth, beginning of exponential phase, midexponential phase and beginning of stationary phase. Lignin-amended cultures achieved twice the cell biomass as unamended cultures over three days, and in this time degraded 60% of lignin. Transcripts in early exponential phase reflected this accelerated growth. A complement of laccases, aryl-alcohol dehydrogenases, and peroxidases were most up-regulated in lignin amended conditions in mid-exponential and early stationary phases compared to unamended growth. The association of hydrogen production by way of the formate hydrogenlyase complex with lignin degradation suggests a possible value added to lignin degradation in the future.

Orellana, Roberto (ORCID:0000000306333458)↗
DOE OSTI2019

Stress response to CO 2 deprivation by Arabidopsis thaliana in plant cultures

After being the standard plant propagation protocol for decades, cultures of Arabidopsis thaliana sealed with Parafilm remain common today out of practicality, habit, or necessity (as in co-cultures with microorganisms). Regardless of concerns over the aeration of these cultures, no investigation has explored the CO 2 transport inside these cultures and its effect on the plants. Thereby, it was impossible to assess whether Parafilm-seals used today or in thousands of older papers in the literature constitute a treatment, and whether this treatment could potentially affect the study of other treatments.For the first time we report the CO 2 concentrations in Parafilm-sealed cultures of A. thaliana with a 1 minute temporal resolution, and the transcriptome comparison with aerated cultures. The data show significant CO 2 deprivation to the plants, a drastic suppression of photosynthesis, respiration, starch accumulation, chlorophyll biosynthesis, and an increased accumulation of reactive oxygen species. Most importantly, CO 2 deprivation occurs as soon as the cotyledons emerge. Gene expression analysis indicates a significant alteration of 35% of the pathways when compared to aerated cultures, especially in stress response and secondary metabolism processes. On the other hand, the observed increase in the production of glucosinolates and flavonoids suggests intriguing possibilities for CO 2 deprivation as an organic biofortification treatment in high-value crops.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Oligosaccharide production and signaling correlate with delayed flowering in an Arabidopsis genotype grown and selected in high [CO 2 ]

Since industrialization began, atmospheric CO 2 ([CO 2 ]) has increased from 270 to 415 ppm and is projected to reach 800–1000 ppm this century. Some Arabidopsis thaliana (Arabidopsis) genotypes delayed flowering in elevated [CO 2 ] relative to current [CO 2 ], while others showed no change or accelerations. To predict genotype-specific flowering behaviors, we must understand the mechanisms driving flowering response to rising [CO 2 ]. [CO 2 ] changes alter photosynthesis and carbohydrates in plants. Plants sense carbohydrate levels, and exogenous carbohydrate application influences flowering time and flowering transcript levels. We asked how organismal changes in carbohydrates and transcription correlate with changes in flowering time under elevated [CO 2 ]. We used a genotype (SG) of Arabidopsis that was selected for high fitness at elevated [CO 2 ] (700 ppm). SG delays flowering under elevated [CO 2 ] (700 ppm) relative to current [CO 2 ] (400 ppm). We compared SG to a closely related control genotype (CG) that shows no [CO 2 ]-induced flowering change. We compared metabolomic and transcriptomic profiles in these genotypes at current and elevated [CO 2 ] to assess correlations with flowering in these conditions. While both genotypes altered carbohydrates in response to elevated [CO 2 ], SG had higher levels of sucrose than CG and showed a stronger increase in glucose and fructose in elevated [CO 2 ]. Both genotypes demonstrated transcriptional changes, with CG increasing genes related to fructose 1,6-bisphosphate breakdown, amino acid synthesis, and secondary metabolites; and SG decreasing genes related to starch and sugar metabolism, but increasing genes involved in oligosaccharide production and sugar modifications. Genes associated with flowering regulation within the photoperiod, vernalization, and meristem identity pathways were altered in these genotypes. Elevated [CO 2 ] may alter carbohydrates to influence transcription in both genotypes and delayed flowering in SG. Changes in the oligosaccharide pool may contribute to delayed flowering in SG. This work extends the literature exploring genotypic-specific flowering responses to elevated [CO 2 ].

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

The RNA-binding protein Modulo promotes neural stem cell maintenance in Drosophila

A small population of stem cells in the developing Drosophila central nervous system generates the large number of different cell types that make up the adult brain. To achieve this, these neural stem cells (neuroblasts, NBs) divide asymmetrically to produce non-identical daughter cells. The balance between stem cell self-renewal and neural differentiation is regulated by various cellular machinery, including transcription factors, chromatin remodelers, and RNA-binding proteins. The list of these components remains incomplete, and the mechanisms regulating their function are not fully understood, however. Here, we identify a role for the RNA-binding protein Modulo (Mod; nucleolin in humans) in NB maintenance. We employ transcriptomic analyses to identify RNA targets of Mod and assess changes in global gene expression following its knockdown, results of which suggest a link with notable proneural genes and those essential for neurogenesis. Mod is expressed in larval brains and its loss leads to a significant decrease in the number of central brain NBs. Stem cells that remain lack expression of key NB identity factors and exhibit cell proliferation defects. Mechanistically, our analysis suggests these deficiencies arise at least in part from altered cell cycle progression, with a proportion of NBs arresting prior to mitosis. Overall, our data show that Mod function is essential for neural stem cell maintenance during neurogenesis.

Parra, Amalia S.↗
DOE OSTI2013

The Toxoplasma gondii cyst wall protein CST1 is critical for cyst wall integrity and promotes bradyzoite persistence

Toxoplasma gondii infects up to one third of the world’s population. A key to the success of T.gondii is its ability to persist for the life of its host as bradyzoites within tissue cysts. The glycosylated cyst wall is the key structural feature that facilitates persistence and oral transmission of this parasite. We have identified CST1 (TGME49_064660) as a 250 kDa SRS (SAG1 related sequence) domain protein with a large mucin-like domain. CST1 is responsible for the Dolichos biflorus Agglutinin (DBA) lectin binding characteristic of T. gondii cysts. Deletion of CST1 results in a fragile brain cyst phenotype revealed by a thinning and disruption of the underlying region of the cyst wall. These defects are reversed by complementation of CST1. Additional complementation experiments demonstrate that the CST1-mucin domain is necessary for the formation of a normal cyst wall structure, the ability of the cyst to resist mechanical stress and binding of DBA to the cyst wall. RNA-seq transcriptome analysis demonstrated dysregulation of bradyzoite genes within the various cst1 mutants. These results indicate that CST1 functions as a key structural component that reinforces the cyst wall structure and confers essential sturdiness to the T. gondii tissue cyst.

bradyzoite↗
DOE OSTI2020

Single cell heterogeneity in influenza A virus gene expression shapes the innate antiviral response to infection

Viral infection outcomes are governed by the complex and dynamic interplay between the infecting virus population and the host response. It is increasingly clear that both viral and host cell populations are highly heterogeneous, but little is known about how this heterogeneity influences infection dynamics or viral pathogenicity. To dissect the interactions between influenza A virus (IAV) and host cell heterogeneity, we examined the combined host and viral transcriptomes of thousands of individual cells, each infected with a single IAV virion. We observed complex patterns of viral gene expression and the existence of multiple distinct host transcriptional responses to infection at the single cell level. We show that human H1N1 and H3N2 strains differ significantly in patterns of both viral and host anti-viral gene transcriptional heterogeneity at the single cell level. Our analyses also reveal that semi-infectious particles that fail to express the viral NS can play a dominant role in triggering the innate anti-viral response to infection. Altogether, these data reveal how patterns of viral population heterogeneity can serve as a major determinant of antiviral gene activation.

59 BASIC BIOLOGICAL SCIENCES↗